In the United States today, transfusion of incompatible blood is a significant factor contributing to morbidity and mortality especially for individuals whose clinical condition or disease—such as sickle cell disease, thalassemia and hematologic malignancies including leukemia and myelodysplastic syndrome—requires periodic transfusion of one or multiple types of blood cells. At present, a minimal degree of compatibility between donor and recipient “blood types” is ascertained in accordance with a “type & screen” paradigm by determining the major phenotypes, defined in terms of the antigens AB (which define the ABO blood groups) and RhD, and screening recipients for alloantibodies against other antigens, and only if such antibodies are detected identifying the antibody, or antibodies, in order to select donor cells lacking the corresponding antigen(s) (“antigen-negative blood”) (Hillyer, C. D. et al., Blood Banking and Transfusion Medicine: Basic Principles & Practice, Elsevier Science Health Science 2002). The repertoire of serologic testing methodologies for addressing this task include: direct agglutination, immediate spin test, as well as indirect antiglobulin test (referred to as “IAT”; see I. Dunsford et al., Techniques in Blood Grouping, 2nd ed. Oliver and Boyd, Edinburgh (1967)). The IAT detects antibodies in the recipient's plasma that recognize one of the major antigens (A, B and RhD) expressed on a donor's erythrocytes which thereby can elicit an adverse transfusion reaction.
Reactions may vary in severity ranging from “none” to “severe” (Hillyer, C. D. et al., supra at p. 17). For instance, critical antigens in the ABO or Rh blood groups, if mismatched, can induce a severe adverse reaction, whereas antigen N, if mismatched, does not. The degree of severity also varies depending upon whether the subject is an adult or a newborn child. For example, an offending antigen S may cause only a mild adverse reaction in an adult but can cause severe hemolytic disease of the newborn. A primary clinical concern arises from the fact that transfusion reactions may entail the accelerated destruction of administered cells, thereby diminishing the efficacy of the therapeutic intervention. Thus, hematology patients undergoing chemotherapy often develop pancytopenias that, while initially caused by impaired hematopoieisis, often are exacerbated by antibody-mediated destruction of incompatible cells given during treatment; a case in point is thrombocytopenia: as patients become increasingly less responsive and not infrequently completely unresponsive (“refractory”) to platelet transfusion, they are exposed to an increased risk of bleeding which may be fatal. To mitigate this risk, these patients receive more frequent transfusions to maintain at least a minimal platelet count, resulting in excess consumption of product; these patient also often require extended care in hospital, at significant excess expense (Meehan et al 2000, “Platelet Refractoriness: Utilization and Associated Costs in a Tertiary Care Hospital”, American J of Hematology, 64: 251-256 (2000)).
Reducing the risk of allo-immunization, by preventing the exposure of the patient to unacceptably immunogenic epitopes, thus remains an important clinical concern, especially in the context of the emphasis on patient-centric and preventive medicine codified in the Affordable Care Act, Health and Human services website, at healthcare/rights/law/index.html).
The current practice of confirming the compatibility of units intended for transfusion for only the major antigens and, in the event, for additional antigens if corresponding specific antibodies are detected, is ill suited to advance this objective. In fact, this “re-active” approach often exacerbates the problem when patients require multiple transfusions. This is so because “antigen-negative” units invariably expose these patients to new allo-antigens and trigger the proliferation of antibodies whose identification in turn requires increasingly complex laboratory procedures followed by the search, often under time pressure, for increasingly less common donor units that lack all the cognate antigens for the antibodies now in the mix: a sisyphean task, with undesirable consequences for patients, who are needlessly exposed to increased clinical risk, and for payers, who bear the excess cost.
The re-active approach, in part, reflects the limitations of the available repertoire of serologic methods which are effective only when antibodies are already manifest. In addition, the extension of routine serologic typing to all clinically relevant antigens is precluded by the lack of appropriate reagents including antisera and the complexity and limited reliability of labor-intensive protocols requiring special expertise and training in immunohematology, particularly when encountering multiple alloantibodies or weakly expressed antigens. Sensitivity is another concern regarding the accuracy of the results, serotyping is based on the interpretation of agglutination patterns that reflect the level of antigen expression, and weakly expressed antigens may be missed.
In contrast, modern methods of DNA analysis such as those described in “BeadChip Molecular Immunohematology” (see Amazon's website under “BeadChip-Molecular-Immunohematology-Profiling-Analysis/dp/144197511X”) are free from these limitations. Thus, the analysis of blood group genes at the DNA level provides a detailed picture of the allelic diversity that underlies phenotypic variability. As described in a number of sources (including, Hashmi et al., Transfusion, 45, 680-688 (2005)) available methodologies permit the simultaneous analysis of clinically significant single nucleotide polymorphisms within the genes encoding antigens in the Kell, Duffy, Kidd, MNS and other systems including the highly variable RhD and RhCE genes, Human Leukocyte Antigens, Human Platelet Antigens and others. See U.S. Pat. No. 7,612,193 (incorporated by reference). Furthermore, these methods can eliminate the need for costly reagents and complex and labor-intensive protocols for serologic analysis, as well as the need for repeat testing of recipients for antibodies to particular donor antigens, and help in addressing clinical problems that cannot be addressed by serologic techniques, such as: the determination of antigens for which the available antibodies are only weakly reactive; the analysis of recently transfused patients; or the identification of fetuses at risk for hemolytic disease of the newborn.
The benefit of identifying immunogenic epitopes on the basis of genotypes relating to the expression of transfusion antigens is to minimize or eliminate not only the risk of antibody proliferation with its adverse effects, but also the risk of immunizing recipients in the first place, and to enable the rapid selection of blood products for transfusion from a group of donors.
Thus, to reduce the risk of allo-immunization, and the antibody-mediated accelerated destruction of administered cells, it will be preferable to pro-actively align molecular signatures of recipient and candidate donor(s) by deploying modern methods of DNA analysis.
While DNA analysis is the de facto standard for identifying stem cell and solid organ donors, the prediction of (T-cell receptor) epitope configurations as a function of differences in DNA sequence between donor and recipient has remained a challenging and elusive task in that setting. Currently, tissue banks seek graft donors whose Human Leukocyte Antigen (“HLA”) alleles must match the recipient's at multiple class I and class II loci to within not more than two mismatched positions to be acceptable. Thus, donors are assessed on the basis of sequence similarity, such that, when a perfect match is not available, graft selection is guided by mere rules of thumb that may deprive certain recipients of perfectly acceptable (stem cell as well as solid organ) transplants. A rational basis for assessing the impact of mismatched alleles on epitope configurations and the associated risk of an adverse reaction (such as graft rejection or graft-vs-host disease) remains to be established. (S. Feng, Characteristics associated with Liver Graft Failure: The Concept of a Donor Risk Index,” Am. J. of Transplantation, Vol. 6, pp. 783-790 (2006); K. Lentine, “Cardiovascular Risk Assessment Among Potential Kidney Transplant Candidates: Approaches and Controversies” Am. J. of Kidney Diseases, Vol. 55, pp. 152-167 (2010).). In contrast, the use of DNA analysis, in the process described herein, forms a critical input to the exclusion of donor units that would expose the intended recipient to unacceptably immunogenic epitopes. Thus, the process described here differs from the genetic cross-matching currently used by tissue banks for the selection of stem cells for allogeneic transplants.
A problem in using DNA analysis for establishing molecular signatures is how to store genetic information in such a way as to permit the selection of suitable products for patients, especially when patients do not receive all treatment at the same institution: this often is the case with sickle cell patients in crisis, or hematology patients commencing treatment in the community setting who ultimately require tertiary care at another institution. The genetic information is more complex than is information obtained by serotyping, and the capability of interpreting it providing it in clinically actionable form is less widely available. An additional problem with genetic analysis is that, as new markers become known, their clinical significance must be assessed so that new markers, if significant, can be added to the analysis guiding the identification of compatible products for a patient. Accordingly, a system of storing and keeping current such information, and for allowing secure access to it for providing compatible products to a patient is needed.
Thus, to enable a pro-active approach to the selection of appropriate cells for transfusion, a viable process is needed by which to generate, make available to the patient in “portable” and preferably in “wearable” form, and maintain up-to-date the relevant clinical information provided by these methods while maintaining the confidentiality of the patient's personal information.